Aedes aegypti mosquitoes are the principal vectors for several human diseases including Dengue Fever, which generates ~50 million cases per year and places 2.5 billion people at risk (17). Novel strategies to combat mosquito-borne diseases are in high demand for A. aegypti as well as other mosquitoes such as the malarial vector Anopheles gambiae. Our goal is to discover new ways to interfere with the mosquito's ability to locate a human host for a blood meal. However, the mechanistic basis of host-seeking and its regulation remains incompletely understood. Although it is known that mosquitoes require human odor cues to locate a human host, the critical odor components and associated olfactory receptors have not been identified (12, 23). Mosquito host-seeking behavior is inhibited for three days following a blood meal. This suppression of feeding behavior is initiated in part by the release of the neuropeptide Head Peptide-I, but the mechanism by which this peptide alters mosquito behavior and the receptor through which it signals is unknown (4). One possible mechanism by which Head Peptide-I may inhibit host-seeking behavior is to modulate the mosquito's ability to smell a human host. To investigate this hypothesis, we propose to identify the Head Peptide-I responsive neurons by de-orphanizing and localizing the currently unknown Head Peptide-I receptor. Using a cell-based calcium-imaging screen, we have identified the G-protein coupled receptor NPY-Like Receptor-1 (NPYLR1) as a candidate Head Peptide-I receptor. Supporting data reveals that multiple NPYLR1 agonists are capable of inhibiting host-seeking behavior when injected into non-bloodfed females. To investigate whether NPYLR1 is required for Head Peptide-I inhibition, we are using recently pioneered targeted mutagenesis with zinc-finger nucleases to create NPYLR1 null-mutant mosquitoes. We predict that these mutants will no longer lose responsiveness to human odor cues after a blood meal. To localize Head Peptide-I responsive NPYLR1 neurons, we will use a custom NPYLR1 antibody and create transgenic mosquitoes that fluorescently mark NPYLR1 expressing cells. This information will clarify the neural mechanism of Head Peptide-I inhibition, and could form the basis of novel strategies to control mosquito host-seeking behavior. PUBLIC HEALTH RELEVANCE: Aedes aegypti mosquitoes are the principal vectors of several human diseases including Dengue Fever, which places 2.5 billion people worldwide at risk and produces ~50 million cases per year (17). Novel strategies to combat mosquito-borne diseases in A. aegypti as well as other mosquitoes, like the malaria vector Anopheles gambiae, are in high demand. The research in this application aims to define a neuromodulatory pathway that controls host-seeking behavior in mosquitoes that will identify novel targets for disrupting their ability to locate and feed on human hosts.